The promoter of the HaCaT keratinocyte differentiation-related gene keratin 4 contains a functional AP-2 binding site.
The nuclear transcription factor AP-2 appears to be a key regulator mediating programmed gene expression during embryonic morphogenesis and adult cell differentiation. AP-2 has also been considered to be involved in epidermal gene regulation, but its precise role is not yet defined. The level of AP-2 transcripts increases during culturing of HaCaT keratinocytes preceding the expression of the differentiation-related gene keratin 4 (K4). The current study was aimed at investigating whether AP-2 transactivates K4 transcription. We cloned and sequenced the promoter region of K4 and found, in addition to canonical sequences, an AP-2 consensus site in the vicinity of the transcriptional start. In order to provide functional evidence for a regulation of K4 transcription by AP-2, we cloned various parts, which did or did not contain the AP-2 site of the K4 upstream sequence, into Cat reporter plasmids. These constructs were ballistically transfected into differentiating HaCaT keratinocytes. The determination of the resulting Cat activity revealed that the AP-2 site in the vicinity of the transcriptional start was functional for K4 transcription. Thus, the role of AP-2 in the process of keratinocyte differentiation appears to be considerable. In addition, further regulatory elements were found to be necessary for full transcription of K4.
Wanner
R
R
Institute of Molecular Biology and Biochemistry, UKBF, FU Berlin, Berlin, Germany. rw@zedat.fu-berlin.de
Zhang
J
J
Dorbic
T
T
Mischke
D
D
Henz
B M
BM
Wittig
B
B
Rosenbach
T
T
D001483:Base Sequence; D001665:Binding Sites; D002454:Cell Differentiation / Q000235:genetics; D002460:Cell Line; D003001:Cloning, Molecular; D016384:Consensus Sequence; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015870:Gene Expression; D017930:Genes, Reporter; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism*; D007633:Keratins / Q000235:genetics*; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection
D004268:DNA-Binding Proteins; D050656:Transcription Factor AP-2; D014157:Transcription Factors; D007633:Keratins